Kocatepe Veterinary Journal

Kocatepe Vet J (2020) 13(3):294-303 RESEARCH ARTICLE DOI: 10.30607/kvj.728554

The Relationship Between Profile and Expression Levels of SCD, FASN and SREBPF1 Genes in Damascus Dairy Goats

Hüseyin ÖZKAN1*, Akın YAKAN1

1Hatay Mustafa Kemal University, Faculty of Veterinary Medicine, Department of Genetic, Hatay, Turkey

ABSTRACT In this study, the relationship between the expression levels of SCD, FASN, SREBPF1 genes and milk fatty acid profiles in goat with low (LSFA) and high (HSFA) saturated fatty acid content was investigated and correlated. In HSFA group, SCD, FASN and SREBPF1 genes were approximately 7, 9 and 4 folds more expressed than LSFA in milk somatic cells, respectively (P<0.01). Also, positive correlations were determined between SCD and FASN (0.907; P<0.001), SCD and SREBPF1 (0.628; P<0.001), FASN and SREBPF1 (0.720; P <0.001) genes. Positive and important correlation was found between C6:0 () and SCD (0.468; P<0.05) and SREBPF1 (0.388; P<0.05) genes. On the other hand, positive correlations were found between all of these three genes and C8:0 () and C10:0 (). In milk samples, C14:0 () and SREBPF1 genes were correlated positively (0.469; P<0.05). Moreover, positive correlation was found between the odour index and SCD (0.553; P<0.01), FASN (0.444; P<0.05), SREBPF1 (0.499, P<0.05) genes. The results showed that SCD, FASN and SREBPF1 genes have important effects on the milk fatty acid profile and milk quality of goats and these genes may be candidate in selection applications in goats. Keywords: Gene expression, Goat, Milk fatty acids, Somatic cells ***

Damascus Keçilerinde Süt Yağ Asidi Profili ile SCD, FASN ve SREBPF1 Genlerinin Ekspresyon Seviyeleri Arasındaki İlişki

ÖZ Bu çalışmada, doymuş yağ asidi içeriği düşük (LSFA) ve yüksek (HSFA) olan keçi sütlerinde SCD, FASN, SREBPF1 genlerinin ekspresyon düzeyleri ile süt yağ asidi profilleri ve aralarındaki ilişki araştırılmıştır. HSFA grubunda süt somatik hücrelerinde SCD, FASN ve SREBPF1 genleri, LSFA grubuna göre yaklaşık sırasıyla 7, 9 ve 4 kat daha fazla ifade edilmiştir (P<0,01). Ayrıca, SCD ve FASN (0,907; P<0,001), SCD ve SREBPF1 (0,628; P<0,001), FASN ve SREBPF1 (0,720; P <0,001) genleri arasında pozitif korelasyonlar tespit edilmiştir. C6:0 (Kaproik asit) ile SCD (0,468; P<0,05) ve SREBPF1 (0,388; P<0,05) genleri arasında pozitif ve önemli korelasyonlar bulunmuştur. Bununla birlikte, bu üç genin tamamı ile C8:0 (Kaprik asit) ve C10:0 (Kaprilik asit) yağ asitleri arasında pozitif korelasyonlar belirlenmiştir. Süt örneklerinde C14:0 (Miristik asit) ve SREBPF1 geni arasında pozitif korelasyon gözlenmiştir (0,469; P<0,05). Ayrıca, koku indeksi ile SCD (0,553; P<0,01), FASN (0,444; P<0,05), SREBPF1 (0,499, P<0,05) genleri arasında pozitif korelasyon tespit edilmiştir. Sonuçlar, SCD, FASN ve SREBPF1 genlerinin keçilerin süt yağ asidi profili ve süt kalitesi üzerinde önemli etkileri olduğunu ve bu genlerin keçilerde seleksiyon uygulamalarında aday olabileceğini göstermiştir. Anahtar Kelimeler: Gen ekspresyonu, Keçi, Süt yağ asitleri, Somatik hücre

To cite this article: Özkan H. Yakan A. The Relationship Between Milk Fatty Acid Profile and Expression Levels of SCD, FASN and SREBPF1 Genes in Damascus Dairy Goats. Kocatepe Vet J. (2020) 13(2):294-303

Submission: 28.04.2020 Accepted: 20.07.2020 Published Online: 05.09.2020 ORCID ID; HÖ: 0000-0001-5753-8985, AY: 0000-0002-9248-828X *Corresponding author e-mail: [email protected]

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fatty acids triggers cardiovascular diseases; however INTRODUCTION polyunsaturated fatty acids (PUFA) are protective against cardiovascular diseases (Simopoulos et al. The basic principle of animal breeding is to obtain 2008). On the other hand, it has been stated that healthy populations with superior characteristics in subacute ruminal acidosis may be estimated in terms of yield and quality. With this principle, ruminants with the fatty acid profile in milk, and milk sustainable breeding studies continue in animal fatty acids may be used as biomarkers for detecting husbandry. Researchers have focused on characters subacute ruminal acidosis (Colman et al. 2015, such as meat yield, milk yield and milk quality in Yurchenko et al. 2018). ruminants, which are important farm animals In goats, milk secretion type is apocrine. (Akcapınar and Ozbeyaz 1999, Ozkan and Yakan Therefore, the number and content of somatic cells in 2017). Increasing awareness of nutrition raises the milk are different. The somatic cells that provide the demand for healthy and quality food. The world natural defense mechanism of the mammary gland are population is increasing day by day. Currently, the composed of leukocytes, polymorphous nuclear cells, population of around 8 billion people is expected to and largely epithelial cells. In ruminants, somatic cell be around 10 billion by 2050 (Anonymous 2020a). count is one of the most important parameter that Increasing population and consumption awareness gives information about mammary health and animal requires higher quality production in animal foods physiology. Molecular activity in milk somatic cells is such as eggs, meat, milk and dairy products (Haile et reported to be an important marker for al. 2016, Balthazar et al. 2017). understanding about biological activity in the The World Health Organization (WHO) states that mammary gland (Murrieta et al. 2006, Feng et al. the saturated/unsaturated fatty acids and the n6/n3 2007). In molecular studies related to mammary ratios should be less than 1.0 and 4.0, respectively health and activity, it is possible to study from milk (WHO, 2013). Studies show that goat milk has more somatic cells without the need invasive methods such positive values in terms of these values than cow and as biopsy in tissues (Feng et al. 2007, Jacobs et al. sheep milk (Ceballos et al. 2009, Silanikove et al. 2013). In addition to giving ideas about mammary 2010, Yakan et al. 2019). On the other hand, goat health, somatic cells reflect the biological activity in milk is more digestible than cow’s milk. The goat molecular pathways such as lipogenesis in mammary milk’s fat globules are smaller and also milk is richer tissue (Murrieta et al. 2006). in terms of short and medium chain fatty acids Although there are a lot of researches about (Kompan and Komprej 2012, Yurchenko et al. 2018). the molecular activities of genes in the lipogenesis Goat breeding is an increasingly important form of pathway in non-ruminant species, what is known livestock for some dairy products such as cheese and about the relationship between these genes and milk ice cream (Yurchenko et al. 2018). It is estimated that fat synthesis in ruminants, especially goats, is quite there are more than 1000 goat breeds in the world limited (Yao et al. 2017). The mammary gland in and the total number of goats is around 1 billion lactation is one of the major organs where today (Crisa et al. 2016, Anonymous 2020b). Today, synthesis takes place (Rudolph et al. 2010). It has the number of goats increased approximately 2-folds been reported that SCD (Stearoyl-CoA Desaturase), compared to 10 years ago and it is reported that more along with many factors in goats, is particularly than 10 million goats exist in Turkey (Anonymous effective in the formation of milk fatty acid 2020a). In addition to pure and native breeds such as composition (Kompan and Komprej 2012). SCD is the Hairy goat, Angora goat, cultural breeds such as highly responsible for fatty acid elongation in milk the Saanen, German Alaca Noble goat, Toggenburg (Kompan and Komprej 2012). FASN (Fatty Acid and Damascus goat, and hybrid breeds such as the Synthase) and SREBPF1 (Sterol Regulatory Element- Kilis, Akkeçi, Bornova goat constitute the Turkish Binding Transcription Factor 1) are another goat population. Damascus goat, due to its high milk important regulators of lipogenic pathway. In this yield and fertility properties, is an important animal study, it is aimed to investigate the expression levels breeding in countries such as Turkey, Syria, Lebanon, of SCD, FASN and SREBPF1 genes in the goat milk Israel and Cyprus (Keskin 2002, Yakan et al. 2019). with low and high saturated fatty acid content and to While quality in animal products has been reveal the relationship between the expression levels associated with hygiene and cheating formerly, today of these genes and the milk fatty acid profile in goat it is evaluated in terms of parameters such as milk. composition, processing and technology of the product (Raynal-Ljutovac et al. 2005, Yakan et al. 2019). Milk fatty acid composition is one of the most MATERIALS and METHODS important factors affecting the quality of milk and dairy products. The amounts and rates of saturated Animal Materials and unsaturated fatty acids in milk composition In the study, 20 milk samples were taken from 2-3 considerably affect milk quality (Yakan et al. 2019). It years old Damascus goats, which are in the 4th has been reported that the consumption of saturated month of lactation. Goats were under semi-intensive 295 feeding. Approximately 100 ml milk collected to Saturated Fatty Acid Content (HSFA). The groups nuclease free and sterile tubes from each goat from were created by ordering the samples from large to the morning milking (50 ml volume falcons were used small according to the SFA contents. The first 10 for sample collection). Samples were CMT (California samples constitute LSFA group, while the last 10 Mastitis Test) negative and animals were healthy. samples formed HSFA group. Then, milk samples were quickly transferred to the laboratory for analyzes under cold chain. Total RNA Isolation and cDNA Synthesis Total RNA isolation was performed from milk Cream and Somatic Cell Collection somatic cells according to the TRI-Reagent (Sigma- The samples were centrifuged at 1800 xg for 15 Aldrich, USA) kit protocol (Rio et al. 2010). After minutes at + 4 °C. After centrifugation, samples were chloroform-isopropyl alcohol-ethyl alcohol steps, the kept at – 20 °C for cream layer for 10 min. The cream RNA pellets were obtained from the samples. layer of samples was collected to new tubes with the Depending on their size, the RNA pellets were help of a spatula. Cream samples were kept at – 20 °C diluted with 30-100 µl of nuclease-free water and until fatty acid analyzes. checked for quality. After cream layer was removed, the supernatants in The purity (A260/280) and concentration of the falcon tubes were discarded and the pellets at the RNA isolated from the samples were checked in the bottom of falcons were transferred to 15 ml, sterile nucleic acid meter (Merinton, SMA-1000 UV and nuclease-free falcon tubes. Cell suspensions in Spectrophotometer). Also, 28S and 18S rRNA the falcon tubes were completed with PBS to 15 ml. subunits were evaluated in 1% agarose gel for RNA Samples were centrifuged at 1800 xg for 15 minutes quality controls (100 V and 25 min). at +4 °C. Then, supernatants of samples were poured The DNase treatment was performed to samples for out; 1 ml TRI Reagent (Sigma-Aldrich, USA) solution possible genomic DNA contamination (DNase I, was added to the cell pellets of samples. Total RNA RNase free, Thermo Scientific, USA, Cat no: isolation was made from the cell pellets homogenized EN0525). According to the Revertaid First Strand in TRI Reagent. cDNA Synthesis kit (Thermo-Scientific, USA) protocol, cDNA synthesis was performed. In the Fatty Acid Analysis thermal cycler (Bio-Rad T100, USA), the reaction was Approximately 500 µl cream from each sample was carried out for 10 min at 25 °C, 120 min at 37 °C, and used for fatty acid analysis. The creams were 5 min at 85 °C. After reaction, the final volume of the homogenized with 2 ml of 2N methanolic KOH for samples were completed to 200 µL with nuclease free 4 minutes at room temperature. Thereafter, 4 ml of n- water and kept at – 80 °C until gene expression Heptane (Merck, USA) was added to the analyses were performed. homogenate. After homogenization for 2 minutes at room temperature, samples were centrifuged at 200 xg for 5 minutes to phase separation. Methyl esters Quantitative Real-Time PCR Analysis collected in the upper phase were transferred to vials Amplification of genes were performed using 10 µl with a volume of 1.5 ml and fatty acids were of each cDNA sample with the kit containing SYBR determined by HP Innowax column (60 m length, Green I dye (Power SYBR® Green PCR Master, 0.25 mm i.d. x 0.25 µm film) in Gas Chromatography ThermoFisher Scientific, USA, Cat no: 4367659). The (HP Agilent 6890, USA). Injector temperature was set reaction was arranged 10 minutes at 95 °C, followed at 250 °C and detector temperature at 270 °C. Helium by 15 seconds at 95 °C, 60 seconds at 60 °C, and 40 was used as the carrier gas. Injection was splitless cycles in qPCR (Bio-Rad CFX-96 Touch Real time with a total injection volume of 1 μl and injector was PCR, USA). Each cDNA sample was studied as washed three times with n-Heptan. Oven temperature duplicate and ACTB gene was used as reference gene. was programmed initially at 90 °C for 3 min and was The forward and reverse sequences of the primers increased to 250 °C with a 3 °C/min ramp rate. The used in the amplification of genes are shown in Table determined peaks on chromatogram were verified by 1. comparison of retention times of internal standards (FAME Mix, Supelco, USA) According to the results of fatty acid contents of milks, the samples were divided into two groups as Low Saturated Fatty Acid Content (LSFA) and High

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Table 1. Forward and reverse sequences of primers of studied genes FORWARD AND REVERSE PRODUCT GENES PRIMER SEQUENCES* LENGTH F: 5’-GCACACAATATGGACCCCCA-3’ FASN 183 R: 5’-CATGCTGTAGCCTACGAGGG-3’ F: 5’-ATCGCCCTTACGACAAGACC-3’ SCD 186 R: 5’-CATAAGCCAGACCGATGGCA-3’ F: 5’-AACATCTGTTGGAGCGAGCA-3’ SREBPF1 134 R: 5’- TCCAGCCATATCCGAACAGC-3’ F: 5’-TGGATCGAGCATCCCCAAAG-3’ ACTB 169 R: 5’-ACTGGCCCCTTCTCCTTAGA-3’ *: All primer sequences were designed in this study by the authors via Primer-BLAST (NCBI).

Statistical Analysis and results were given as fold changes (Livak and SPSS package program (Version 23.0) was used to Schimitgen 2001). P<0.05 was accepted as significant. calculate the obtained data. Student-t test was used to measure the difference between the groups. RESULTS Spearman correlation coefficient was used to calculate the correlation between parameters. Odour (Yakan et The saturated fatty acid content was 64.92±0.90(%) al. 2019), thrombogenic and atherogenic indexes were in the LSFA group, while HSFA group saturated fatty calculated using fatty acid parameters (Ulbricht and acid content was 71.27±0.45(%), (P<0.001). Between Southgate 1991). On the other hand, the 2-ΔΔCt groups, variable levels of significance in terms of fatty method was used for gene expression calculations, acids were determined (Table 2).

Table 2. Milk fatty acid profiles of LSFA and HSFA groups (Mean±SEM) Fatty acids (%) LSFA HSFA P C4:0 () 1.62±0.07 1.47±0.15 - C6:0 (Caproic Acid) 2.06±0.08 2.16±0.12 - C8:0 (Caprylic Acid) 2.42±0.11 2.68±0.14 - C10:0 (Capric Acid) 7.60±0,38 9.28±0.33 ** C11:0 (Undecanoic Acid) 0.23±0.02 0.19±0.02 - C12:0 () 2.95±0.17 3.68±0.17 - C14:0 (Myristic Acid) 8.63±0.30 10.63±0.47 ** C14:1 (Miristoleic Acid) 0.44±0.04 0.64±0.31 - C15:0 (Pentadecylic Acid) 1.52±0.09 1.37±0.04 - C15:1 (Pentadecenoic acid) 0.38±0.02 0.31±0.01 * C16:0 () 24.43±0.69 26.7±0.56 * C16:1 () 0.97±0.11 0.82±0.10 - C17:0 () 1.02±0.04 0.87±0.05 ** C17:1 (Heptadecenoic Acid) 0.49±0.03 0.38±0.01 * C18:0 () 11.27±0.47 10.87±0.48 - C18:1 () 26.48±1.13 21.18±0.50 *** C18:2 n6 () 1.46±0.25 1.29±0.20 - C18:3 n3 (Alphalinolenic Acid) 1.57±0.17 1.68±0.17 - C20:0 () 0.71±0.08 0.89±0.20 - C20:1 (Gondoic Acid) 1.42±0.08 0.98±0.04 *** C20:2 n6 (Eicosadienoic Acid) 0.48±0.08 0.41±0.09 - C20:3 n3 (Eicosatrienoic Acid) 0.18±0.02 0.18±0.03 - C20:3 n6 (Dihomo-g –linolenic Acid) 0.13±0.02 0.14±0.02 - C20:4 n6 () 0.18±0.01 0.16±0.01 - C22:0 () 0.38±0.01 0.33±0.01 ** C22:1 () 0.22±0.03 0.16±0.01 - C24:0 () 0.15±0.02 0.19±0.04 - C24:1 () 0.82±0.12 0.64±0.04 - *: P<0.05; **:P<0.01; ***:P<0.001; -:P>0.05

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Differences were found in some parameters nutritional value was lower in LSFA than HSFA determined in samples grouped according to group (P<0.01), atherogenic and thrombogenic index saturated fatty acid content (Table 3). While the values were higher (P<0.001) (Table 3).

Table 3. Some parameters were determined according to the fatty acid compositions in the groups (Mean±SEM) PARAMETERS LSFA HSFA P

ΣSFA 64.92±0.90 71.27±0.45 ***

ΣMUFA 29.79±1.07 24.03±0.56 ***

ΣPUFA 3.87±0.35 3.71±0.28 -

ΣUFA 33.66±0.85 27.74±0.44 ***

Σn6 2.18±0.25 1.87±0.22 -

Σn3 1.70±0.19 1.84±0.17 -

Σn6/n3 1.38±0.17 1.12±0.19 -

OI 13.70±0.60 15.59±0.69 -

NV 1.56±0.08 1.21±0.05 **

AI 1.46±0.07 2.07±0.09 ***

TI 1.27±0.04 1.60±0.03 *** SFA: Saturated fatty acids; MUFA: Monounsaturated fatty acids; PUFA: Polyunsaturated Fatty acids; UFA: Unsaturated fatty acids; OI: Odour index; NV: Nutritional Value; AI: Atherogenic index; TI: Thrombogenic index *: P<0.05; **:P<0.01; ***:P<0.001; -:P>0.05 OI= (C4:0+C6:0+C8:0+C10:0); NV= (C18:0+C18:1)/C16:0; AI = (C12:0+(4*C14:0)+C18:0)/ΣUFA TI= (C14:0+C16:0+C18:0)/((0.5*C18:1)+(0.5*ΣMUFA)+(0.5*Σn6)+(3*Σn3)+(Σn3/Σn6))

Correlation values between milk fatty acid parameters such as SFA, MUFA, PUFA, and UFA in milk are given in Table 4.

Table 4. Correlations between some parameters in milk PARAMETERS ΣMUFA ΣPUFA ΣUFA Σn6 Σn3 Σn6/n3 OI NV AI TI ΣSFA -0.937*** 0.023 -0.997*** -0.119 0.332 -0.364 0.623** -0.820*** 0.929*** 0.889*** ΣMUFA -0.244 0.932*** -0.014 - 0.383* -0.657** 0.815*** -0.892*** -0.789*** 0.523** ΣPUFA 0.005 0.811*** 0.582** 0.212 0.183 -0.194 -0.018 -0.053 ΣUFA 0.156 -0.341 0.398* -0.591** 0.826*** -0.929*** -0.886*** Σn6 0.056 0.683*** 0.147 0.003 -0.198 -0.005 Σn6/n3 -0.023 0.341 -0.442* -0.114 OI -0.361 0.480* 0.411* NV -0.756*** 0.808*** AI 0.847*** *:P<0.05;*:P<0.01; ***:P<0.001

RNA samples had appropriate purity 9 (9.17; P<0.05) and 4 (4.26; P<0.05) folds more in (A260/280=1.87±0.03) and concentration the HSFA group compared to the LSFA group, (283.07±42.37 ng/μL). SCD, FASN and SREBPF1 respectively (Figure 1). genes were expressed approximately 6 (6.35; P<0.05),

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1 5 L S F A *

s H S F A e

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n A

a *

N

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R C

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o F

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S C D F A S N S R E B P F 1

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Figure 1. mRNA fold changes of genes in LSFA and HSFA groups (*: P<0.05)

the monounsaturated fatty acid ratio and the Some correlations were determined between SCD, expression levels of SCD (-0.540; P<0.01), FASN (- FASN, SREBPF1 genes and fatty acid parameters. 0.439; P<0.05) and SREBPF1 (-0.471; P<0.05). In The correlation results showed that there were addition, there was a positive correlation between important, positive and high correlations between SCD gene and polyunsaturated fatty acid ratio (0.426; genes (Between SCD and FASN: 0.907: P<0.001; P<0.05) and negative correlation between the between SCD and SREBPF1: 0.628; P<0.001; unsaturated fatty acid ratio and SCD (-0.433; P<0.01) between FASN and SREBPF1: 0.720: P<0.001). and SREBPF1 (-0.414; P<0.05) were found. But then, Also, with variable levels of significance, positive and a positive correlation was found between the SCD negative correlations were found between genes and and n3 ratio. Odour index was found correlated with fatty acids (Table 5). SCD, FASN and SREBPF1 genes (0.553; 0.444; While there was a positive correlation between 0.499, respectively, P<0.05). A positive correlation saturated fatty acid ratio and SCD (0.444; P<0.05), was found between SREBPF1, a lipogenic FASN (0.344; P>0.05) and SREBPF1 (0.405; transcription factor, and the atherogenic index (0.398; P<0.05), a negative correlation was found between P<0.05).

Table 5. Correlations between genes and fatty acid parameters PARAMETERS& SCD FASN SREBPF1

C6:0 (Caproic Acid) 0.468* 0.370 0.388*

C8:0 (Caprylic Acid) 0.573** 0.457* 0.466*

C10:0 (Capric Acid) 0.492* 0.400* 0.508*

C14:0 (Myristic Acid) 0.197 0.241 0.469*

C17:1 (Heptadecenoic Acid) -0.325 -0.254 -0.432*

C18:1 (Oleic Acid) -0.466* -0.377* -0.478*

C18:3 n3 (Alphalinolenic Acid) 0.460* 0.420* 0.232

C20:1 (Gondoic Acid) -0.385* -0.284 -0.490*

C20:2 n6 (Eicosadienoic Acid) 0.519* 0.339 0.253

ΣSFA 0.444* 0.344 0.405*

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ΣMUFA -0.540** -0.439* -0.471*

ΣPUFA 0.426* 0.259 0.211

ΣUFA -0.433* -0.355 -0.414*

Σn6 0.301 0.099 0.113

Σn3 0.391* 0.356 0.211

Σn6/n3 -0.083 -0.217 -0.087

OI 0.553** 0.444* 0.499*

NV -0.277 -0.140 -0.202

AI 0.287 0.247 0.398*

TI 0.229 0.090 0.223

&: Statistically significant correlations were showed in table *: P<0.05; **: P<0.01

unsaturated fatty acid ratio and it was understood that DISCUSSION this correlation probably occurred due to MUFAs. In HSFA group SCD, FASN and SREBPF1, primary In recent years, with the increasing demand for goat lipogenic genes, were expressed more than 4 folds milk, goat husbandry has gained importance and the compared to the LSFA group and these results level of breeding has increased (Novotna et al. 2019). showed that these genes played an important role in In this study, SCD, FASN and SREBPF1 genes the synthesis of saturated fatty acids in goat mammary expression levels and fatty acid profiles of goats that gland (Figure 1). divided into two groups according to milk SFA SCD has a role as the key regulator in de novo fatty contents were investigated. acid synthesis in mammary gland together with ACC Saturated fatty acids in the diets are associated with and FASN (Bernard et al. 2008). It is reported that cardiovascular diseases (Siri-Tarino et al. 2010). the main function of SCD in the mammary gland Investigation of the factors that play roles in the plays a role in the conversion of long chain fatty formation of variation in dietary fatty acids has acids, especially C16:0 and C18:0, to C16:1 and C18:1, become an important goal. An important part of milk respectively. SCD has a significant effect on the fatty acids in ruminants are saturated fatty acids (Haile fluidity of milk as well (Feng and et al. 2007). In a et al. 2016). One of the most important parameters in study it is reported that the SCD gene is more the formation of milk quality is fatty acid profile expressed during the lactation period in goat (Chilliard et al. 2003). Numerous candidate genes and mammary tissue compared to the dry period, and transcription factors in lipogenic pathways regulate increased SCD expression led to increase of the the formation of milk fatty acid qualities and MUFA content and fat storage in the cell (Yao et al. quantities via production and secretion of fatty acids 2017). In the same study, it is stated that (Moioli et al. 2007, Bionaz and Loor 2008). concentration of oleic acid and synthesis of In HSFA group, while C10:0, C14:0 and C16:0 fatty triacylglycerol are decreased with silencing of SCD acids ratios were more than LSFA group (P<0.01), gene. On the other hand, it is reported in some MUFAs as C15:1, C17:1, C18:1 and C20:1 and C22:0 studies that that animals with high SCD activity in fatty acids ratio were significantly lower (P<0.05). milk may be selected for high quality milk production According to milk fatty acid composition, no that have high UFA content such as oleic acid (Feng significant difference was found between the groups et al. 2007, Jacobs et al. 2013). However, in this study, in terms of other unsaturated fatty acids (Table 2). In while HSFA groups’ SCD gene expression was accordance with the literature, while the atherogenic approximately 6 folds more than LSFA groups and thrombogenic index values were found to be (Figure 1), UFA contents of HSFA groups’ were high in the HSFA group, the unsaturated fatty acid lower than LSFA groups (Table 5). In addition, there ratio and thus the nutritional value were found lower was no difference in oleic acid levels between the (Haile et al. 2016, Idamokoro et al. 2019) (Table 3). groups. It is thought that lactation period and animal As expected, a negative correlation was found race may be caused these results. Since, mammary between the monounsaturated fatty acid ratio and the gland molecular activity and composition of milk odour, atherogenic and thrombogenic indexes. In change in different periods of lactation. On the other addition, it was determined that there was a positive hand, it is reported that the milk fatty acid correlation between the nutritional value and the composition of three different goat breeds were

300 different under similar environmental conditions atherogenic and thrombogenic indexes and SREBPF1 (Idamokoro et al. 2019). (0.469; P<0.05). With these results, it is thought that Abnormal SCD expression in humans is known to be there is a relationship between milk with high associated with obesity and its complications (Huang atherogenic and thrombogenic indexes and et al. 2015, Southam et al. 2015). In cattle and goats, it SREBPF1. is reported that the polymorphism in SCD gene has Significant positive correlations were found between led to change of milk fatty acid composition and target genes of study (SCD, FASN, SREBPF1) and therefore milk quality characteristics have changed C6:0, C8:0, and C10:0 fatty acids (Table 5). SCD, (Conte et al. 2010, Alim et al. 2012). It is reported in a FASN and SREBPF1 genes have important effects study, the over-expression of the SCD gene causes on the synthesis of these fatty acids used in the significant upregulation of some lipogenic genes calculation of the odour index. Capric and caprylic especially as FASN and PPARG in lactating goats acids have been used in medical therapy for (Yao et al. 2017). In this study, it is determined that metabolic-related digestive disorders such as there is a very high correlation between SCD and hyperlipoproteinemia, premature infant feeding, FASN gene (0.907; P<0.001). SREBPF1, which is epilepsy, cystic fibrosis, and coronary heart diseases responsible for the activity of a large number of (Kompan and Komprej 2012). Considering this lipogenic genes such as SCD and FASN in mammals, situation, it is thought that the expression levels of shows a high level of positive correlation with both these genes in the mammary epithelial cells may give SCD (0.628; P<0.001) and FASN (0.720; P<0.001). important ideas in selection applications on milk SREBPF1, a lipogenic transcription factor, is a trigger quality in goats. of SCD and FASN genes and it regulates milk fatty acid profiles with activation in mammary epithelial CONCLUSION cells in lactating ruminants. SREBPF1 is required to trigger SCD expression (Huang et al. 2015, Souhtam In conclusion, the expression levels of SCD, FASN et al. 2015). Disorders in milk and SREBPF1 genes, which are important in the have been reported as a result of abnormal SREBPF1 regulation of energy metabolism, are investigated in activity (Xu et al. 2016). It is reported that SREBPF1 milk with low and high saturated fatty acid content gene expression increased approximately 2 times in and their relationship with fatty acid profile in this cattle, rats and humans, while it increased up to 30 study. It is thought that successful results may be times in goats due to mammary gland physiology in obtained by investigating these genes in the selection the period from pregnancy to lactation (Xu et al. applications on milk quality in ruminants. Also, more 2016). SREBPF1 activity is well defined in humans, studies at the molecular levels are needed to rodents and cattle, but what is known about investigate the importance of fatty acid profile in the SREBPF1 in goats is limited. formation of milk quality. In a study investigating the lipogenic activity in goat breast epithelial cells, SREBPF1 is found to trigger Conflict of Interest: The authors declare that they FASN gene expression (Xu et al. 2016). 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